Since the middle of 1980s, nosocomial infection through ESBL-producing Klebsiella pneumoniae and Escherichia coli has been recognized as a serious problem in Europe and then in U.S.A. Recently, such bacteria tend to gradually increase also in Japan.
ESBLs hydrolyze even broad-spectrum third-generation β-lactam antibacterial agents such as cefotaxime (CTX), ceftazidime (CAZ) and aztreonam (AZT) which are stable against conventional type β-lactamases, and have reduced susceptibility to these medical agents. Continued administration of such medical agents against ESBL-producing bacteria not only would be hopeless from the viewpoint of cure but also might harmfully lead to spreading of ESBL-producing bacteria and developing of new resistant bacteria.
Thus, it is necessary to identify ESBL-producing bacteria through rapid and proper testing and to use proper antibiotics.
Currently, ESBL-detecting testing may be conducted, for example, by 1) method for measuring MICs (minimum inhibitory concentrations) to CTX, CAZ, AZT in the presence and absence of clavulanic acid (CVA), 2) double-disk synergy test method using two kinds of disks, one of which is for CVA and the other of which is for either of CTX, CAZ and AZT, a zone of inhibition around each disk being observed, or 3) E-test method using MIC ratios of CAZ alone and of CAZ with CVA.
However, any of these methods requires MIC measurement through medical-agent susceptibility test to determine the presence of ESBL-producing bacteria, which takes several days for isolation and cultivation of bacteria, actually resulting in failure of rapid determination. Under such circumstances, it has been desired to provide a testing procedure which requires no special operations and devices other than culture of organisms and which is shorter in detection than MIC measurement.
Rapid detection of conventional type β-lactamases such as penicillinase (PCase) and cephalosporinase (CEPase) is concerned with decomposed β-lactam ring of substrate and may be conducted, for example, by (1) acidmetry method for grasping pH change in terms of color change of a pH indicator, (2) iodometry method for utilizing color change in starch-iodine reaction as measure, (3) chromogenic method for grasping change in conjugated system in terms of absorption change in direct visible region and (4) UV method for grasping change in conjugated system in terms of absorption change in ultraviolet region; among these methods, the chromogenic method is said to be most easily accessible from the viewpoint of sensitivity and in that no special devices are required for measurement. Actually, products utilizing chromogenic method with substrate being nitrocefin (JP-56-18197B, U.S. Pat. No. 3,830,700 and British Patent 1408391) are commercially available; however, they react with all β-lactamases and there is no hope of their selective application for ESBL-producing bacteria at all.